Regulatory T cells (Tregs) are critical for actively maintaining immune tolerance. The Foxp family of transcription factors is composed of four members~ Foxp1, Foxp3 and Foxp4 are expressed in lymphocytes. Foxp family members bind DNA as homo-and hetero-dimers to regulate gene expression. Regulatory T cells are CD4+ T cells that express Foxp3. We have generated mice in which both Foxp1 and Foxp4 are deleted in T lymphocytes, leaving Foxp3 the only potentially expressed family member in this T cell subset. Our preliminary data demonstrate that combined loss of Foxp1 and Foxp4 in T lymphocytes substantially alters the development of Tregs in the thymus, reduces peripheral Treg cellularity, and alters suppressive function. Treg development relys on a complex interaction of signals generated by the T cell receptor, costimulatory pathways and cytokine receptors. The overall goal of this proposal is to understand how Foxp1 and Foxp4 alter the development, homeostasis and function of Foxp3+ Tregs. To address this question we propose to generate novel mouse strains in which Foxp1 and Foxp4 are deleted at different stages of Treg development. We will manipulate signaling pathways to determine if the alterations in Foxp1/Foxp4 deficient Tregs are due to altered TCR or cytokine signaling. We will determine if loss of Foxp1 and Foxp4 in Tregs alters in vivo responses in models of transplantation and inflammatory bowel disease. Understanding these fundamental aspects of Treg generation, homeostasis and function are critical to translational strategies of Treg augmentation being developed for use in autoimmune disease and solid-organ transplantation.